Everything you need to know about microbial genetics. Some of the frequently asked questions are as follows:-
Q.1. Name the types of mutations involving many base pairs.
Ans: (1) Deletion (elimination of a region of DNA),
(2) Insertion (addition of new bases to DNA),
(3) Translocation (in which large section of chromosomal DNA gets moved to a new location and in eukaryotes even to a different chromosome),
(4) Inversions (in which the orientation of a particular segment of DNA gets reversed with respect to surrounding DNA).
Q.2. List the rates of mutations of some important types.
Ans: (i) Spontaneous mutations occur at a frequency of 10-6 per generation.
(ii) Transposition events occur more frequently, i.e., at about 10-4
(iii) Nonsense mutations are less frequent, i.e., at 10-6 to 10-8
Q.3. What is the rate of mutation in ribonucleic acid (RNA) genomes? Why is it higher than in DNA genomes?
Ans: The mutation rate in RNA genome is about 1000 mutation is a comparatively short process. The reasons being that the RNA replicates do not seem to have proofreading activities like those of DNA polymerases. Also there a. no comparative RNA repair mechanisms. Instead there are many repair systems for DNA that corrects many a changes before the changes get fixed as mutations.
Q4. What are mutagens?
Ans: Mutagens are chemical, physical or biological agents that enhance the mutation rate by making alterations in DNA. However, the alteration is a mutation only if it is inherited.
Q.5. Name the common chemical mutagens and describe how they cause mutations.
Ans: There are following types of chemical mutagens:
(1) Base analogs:
Many of them are base analogs resembling DNA purine. Pyrimidine bases in structures may consequently lead to faulty pairing, e.g., (1) 5-bromouracil is an analog of thymine. Consequently 5-bromouracil can pair with guanine causing AT to GC substitution. (2) 2-aminopurine serves as an analog of adenine, hence 2- aminopurine can pair with cytosine causing AT to GC substitution.
(2) Alkylating agents:
Alkylating agents such as nitrosoguanidine are powerful mutagens. They may produce direct change even in non replicating DNA. Like base analogs they tend to induce base pair substitutions.
The acridines are intercalating agents leading to microinsertions or microdeletions. The acridines (mutagens) get inserted between DNA base pairs.
Q.6. Which are the physical mutagens?
Ans: Several types of radiations count among physical mutagens.
They are mainly of two types:
(a) Nonionizing radiations, e.g., UV radiations in 260 nm region.
(b) Ionizing radiations: They are short wavelength rays like X-rays, cosmic rays and gamma rays. Among the free radicals the hydroxyl radical (OH) reacts with and inactivates the macromolecules in the cell, of which DNA is the most important.
Q.7. Name the complex cellular mechanism associated with DNA repair.
Ans: SOS regulatory system.
Q.8. What is biological mutagenesis?
Ans: It is the process of transposon mutagenesis. Transposons are widely used by microbial geneticists as mutagenic agents.
Q.9. What is site directed mutagenesis?
Ans: It is the production of specific mutations in specific genes using recombinant DNA technology with the use of synthetic DNA.
Q.10. What is Ames test?
Ans: This test developed under the direction of Bruce Ames at University of California at Berkeley employs a sensitive bacterial assay system for detecting chemical mutagens in the environment. It was primarily conducted for carcinogenic screening.
Q.11. Name the organisms often used in Ames test.
Ans: Histidine auxotroph of Salmonella typhimurium and tryptophan auxotroph’s of Escherichia coli.
Q.12. How does the mechanism of recombination occur in prokaryotes?
Ans: It involves DNA transfer during the process of transformation, transduction and conjugation.
Q.13. Name the protein found in all prokaryotes which assists in pairing required for homologous recombination.
Ans: Rec A protein.
Q.14. What is complementation test?
Ans: Complementation test is used to detect mutations in the same genetic (functional) unit.
Q.15. What is the extreme length of DNA in Bacillus subtilis?
Ans: 1700 pm.
Q.16. Who was the first to show evidence of bacterial transformation?
Ans: British scientist Frederick Griffith in 1928 was the first to prove it with Streptococcus pneumonia (pneumococcus).
Q.17. What were the two strains of S. pneumoniae (pneumococcus) used by F. Griffith and what did he conclude?
Ans: Griffith used two forms, i.e., smooth (virulent) and rough (non-virulent) forms of the bacteria. He found that a mixture of smooth virulent (pathogenic) killed by heat and live, rough non-virulent (nonpathogenic) while injected to mouse proved fatal. It indicated that genetic material (DNA) formed by heat killed smooth type pneumococci brought transformation into the genetic makeup of rough non-virulent form.
Q.18 . What is competence? On what factors does competence depend?
Ans: A cell which can take up a molecule of DNA to be transformed is said to show competence. Competence depends on special proteins and may include membrane associated DNA binding proteins, a cell wall autolysin and various nucleases. In Bacillus subtilize and Streptococcus pneumonia induction of competence depends on the medium and growth stage of the culture.
Q.19. Give variations in uptake of DNA.
Ans: Gram-negative bacterium Haemophilus takes up double-stranded DNA into the cell while in the Gram-positive bacteria Streptococcus and Bacillus only a single DNA strand is taken up while the complementary DNA is simultaneously degraded. However, generally double stranded DNA binds more effectively to the cells. Competent cells may bind 1000 times more DNA than non-competent cells.
Q.20. Give the mechanisms of transformation (Integration of transforming DNA).
Ans: During the process of transformation competent bacteria first bind DNA reversibly, soon however, making it irreversible. The DNA undergoing transformation is bound at the surface of bacterial cells by a DNA binding protein which remains attached to DNA perhaps to prevent it from the attack of enzyme nuclease till the time it reaches the chromosome of the bacterial cell where Rec A protein takes over. It would be worthwhile to add that once the transforming DNA is bound at the surface by the DNA binding protein either the entire double stranded fragment is taken up or one strand is degraded by a nuclease and the other strand is taken up.
Q.21. What is transfection? What is its use?
Ans: It is the transformation of bacteria with DNA extracted from a bacterial virus instead of taking DNA from another bacterium. Transfection is a very useful tool in studying the mechanisms of transformation and recombination as the small size of phase genomes allows for the isolation of a nearly homogenous population of DNA molecules.
Q.22. What do you mean by the term ‘transfection’ in relation to eukaryotes?
Ans: Eukaryotic microorganisms and animal and plant cells can take up DNA in a process which resembles bacterial transformation. Because the word transformation in mammalian cells is used to describe the conversion of cells to the malignant state. Therefore, introduction of DNA into mammalian cells has been called transfection.
Q.23. Name the bacterial genera which possess high efficiency natural transformation.
Ans: High efficiency natural transformation occurs in Azotobacter, Bacillus, Streptococcus, Haemophilus, Neisseria and Thennus, which are easily transformed
Q.24. How can artificial competence be induced in Gram-negative bacterium Escherichia coli?
Ans: Competence can be introduced in E. coli by treating it with high concentration of calcium ions and then stored in the cold. On doing so it becomes transformable at low efficiency. Escherichia coli treated in this way takes up double stranded DNA and therefore, transformation by plasmid DNA is more efficient because no recombination is required. How does calcium treatment work is not well understood but this method is also used with some other Gram-negative bacteria.
Q.25. How is DNA transferred by electroporation?
Ans: Small pores are formed on the membranes of bacterial cells exposed to pulsed electric fields. The DNA molecules present outside the cells during electric pulse, can enter the cells through these pores. By this method plasmid can be directly transferred from one cell to another if both the types of cells are present during electroporation.
Q.26. How can yeast, algae, plant cells, mitochondria and chloroplasts be transfected by particle gun?
Ans: A small steel cylinder containing a gun-powder charge is used to fire nucleic acid coated particles at the target cells. The nucleic acid enters the cell by piercing the wall without damaging the cell and combines with the host DNA.
Q.27. Which are the two main types of transduction?
Ans: Transduction is of two types:
(1) Generalized transduction.
(2) Specialized transduction.
Q.28. What is an episome?
Ans: Plasmids having the ability to integrate into host chromosomes are known as episomes.
Q.29. What is the main mechanism of cell to cell transfer of plasmids?
Q.30. What are conjugative plasmids? Name the gene that controls it.
Ans: Plasmids which govern their own transfer by cell to cell contact are called conjugative plasmids. The set of genes called tra region that controls the transmission of the plasmids.
Q.31. Some plasmids with gene function other than replication not known, have been detected by gel electrophoresis of cell extracts.
Ans: Cryptic plasmids.